Adenoviruses are simple DNA-containing viruses (i.e., composed of only DNA and protein) that multiply in the cell nucleus of the host. These viruses induce latent or acute infections in tonsils, adenoids, lungs, bladder and cornea as well as the gastrointestinal tract and are readily activated. Several adenoviruses are the first common viruses of humans shown to be oncogenic for lower animals under special experimental circumstances. The adenoviruses may serve as "helpers" for adeno-associated viruses which cannot replicate in their absence.
The viral particles of the adenovirus have a dense central core and an outer coat known as the capsid. These particles have an icosahedral configuration and are composed of 252 capsomers: 240 hexons make up the faces and edges of the equilateral triangles and 12 pentons comprise the vertices. The hexons are truncated triangular or polygonal prisms with a central hole. The pentons are more complex, consisting of a polygonal base with an attached fiber protein, whose length (i.e., short or long) varies with viral type. Minor capsid proteins are also associated with the hexons or pentons and confer stability on the capsid to form links with the core proteins, and to function in virion assembly.
Each virion contains one linear, double-stranded DNA molecule associated with proteins to form the core of the adenovirus.
The early region 3 (E3) of adenoviruses plays a critical role in pathogenesis of the virus's disease process even though none of its gene products are essential for replication of the virus in cell cultures. Not all proteins coded in the E3 regions of adenoviruses have been identified, even for the most commonly studied adenovirus, type 2 (Ad2). However, it has been postulated that they mediate cellular or immunological responses through structural or functional homology to regulatory molecules. For this reason, it is possible that proteins generated from the E3 region, or their derivatives, can be used in therapy as modulators of the immune response (e.g., as an immunostimulation system in AIDS patients) or as anti-cancer agents to modify the action of various growth factors. In addition, specific E3 proteins can be used to distinguish between different adenovirus types.
Adenoviruses are widespread in nature. The 89 accepted members of the adenovirus family have similar chemical and physical characteristics and a family cross-reactive antigen but are distinguished by antibodies to their individual type-specific antigens: at least 41 are from humans and the rest from various animals.
The enteric adenoviruses, such as Adenovirus Type 40 or 41 (and also known as Type F Enteric Adenoviruses), are a virus group that cause serious intestinal and diarrheal diseases of young children. In 1978, the World Health Organization initiated a program for global prevention and control for such childhood diseases. As a result, the relative importance of various pathogens in the etiology of diarrhea in many parts of the world has been recognized. For example, rotaviruses, which rank as the most prevalent viral pathogen in childhood diarrhea, may now be close to control as many vaccines are now in sight. This has been made possible through very intensive research over the past decade.
However, the control of enteric adenoviruses, which are responsible for at least 15% of all cases of severe infantile gastroenteritis, is not yet within reach. Although they are second after rotaviruses as viral agents causing this type of infection, enteric adenoviruses remain a poorly defined group of viruses. The paucity of research done on enteric adenoviruses is mainly due to the difficulty of propagating the viruses in cultures. For this reason, there is no sensitive, fast, and diagnostic procedure able to distinguish between enteric adenoviruses and other adenoviruses (Group A, B, C, D, and E) which are commonly present in stools but are not agents of gastroenteritis. Another reason for studying enteric adenoviruses is their possible link to intestinal cancer which appears later in the life of infected individuals.
The standard reference methods for diagnosis of enteric adenoviruses have been (1) immunoelectron microscopy; (2) type-specific neutralization; (3) growth differences on primary human and Graham-293 cells. None of these methods are accurate and suitable for rapid routine use. Recently a new commercially available enzyme-linked immunoabsorbent assay (ELISA) to detect enteric adenoviruses (Adeno-Type 40/41 EIA, Cambridge Bioscience) based on a polyclonal antibody to enteric adenovirus hexon protein was created, but this kit lacks both specificity and sensitivity.
However, the present invention solves the problems associated with the previous methodologies. The present invention describes a recombinant DNA molecule which can produce at least one of Human Adenovirus Type 41 Tak (Ad41) short fiber protein, long fiber protein, or proteins RL-1 to RL-6 of the Ad41 E3 region. (There are presently several isolates known of human adenovirus type 41, but the most common isolate of this adenovirus is human adenovirus type 41 Tak, represented in the present invention. This isolate is the standard Ad41 strain and it is listed in the American Type Culture collection under catalog number ATCC #VR-930.)
The Ad41 short and long fiber protein gene and Ad41 E3 proteins are useful for assays for human enteric adenoviruses since they express only minor immunological cross-reactivity between adenoviruses belonging to different serotypes; they are unique adenovirus proteins (i.e., Ad41 long fiber protein and possibly the short fiber as well are responsible for attachment of the virus to specific cellular receptors in the cell membrane during infection) and they express selective type-specific antigenicity. The genes of the present invention are ideal candidates for specific, selective monoclonal antibodies based on an enzyme immunoassay (EIA) kit, a DNA probe assay system and a vaccine derived from the gene products. The present invention will not only enhance the understanding of the mechanism by which human enteric adenoviruses cause disease in humans, but will also assist in developing molecular probes for diagnosis of such infections.